The present invention relates, in general, to a cancer-related protein and to a nucleic acid sequence encoding same. In particular, the invention relates to a protein over expressed in certain neoplastic cells, including breast and ovarian cancer cells, to its encoding sequence, and to diagnostic and treatment methodologies based on same.
Breast cancer represents the most frequent cause of early morbidity and mortality in women in North America (Harris et al, New Eng. J. Med. 327:319, 390 and 473 (1992)). It is generally believed that this malignancy arises from a multi step process involving mutations in a relatively small number of genes, perhaps 10 or less. These mutations result in significant changes in the growth and differentiation of breast tissue that allow it to grow independent of normal cellular controls, to metastasize, and to escape immune surveillance. The genetic heterogeneity of most breast cancers suggests that they arise by a variety of initiating events and that the characteristics of individual cancers are due to the collective pattern of genetic changes that accumulate (Harris et al, New Eng. J. Med. 327:319, 390 and 473 (1992)).
The classes of genes that are involved in breast cancer are not unlike those found in a number of other well characterized malignancies, although some are highly specific for breast cancer. In particular, mutations in the genes that encode receptors involved in binding to estrogen and progesterone are particularly important because they likely cause the breast cells to proliferate while rendering them unresponsive to the antitumor effects of these hormones in advanced malignancy. In addition, changes in the genes that encode growth factors, other receptors, signal transduction molecules, and transcription factor molecules are frequently involved and have alterations that are involved in the development and progression of breast cancer (King, Nature Genetics 2:125 (1992)). The characterization of the type and number of mutations seen in individual breast cancers is useful in classifying the biological properties of individual cancers and in determining the prognosis for individual patients. For example, the erbB2/HER2/neu gene is particularly valuable in predicting the prognosis of both node-positive and node-negative patients based on the amplification status of the gene (King, Science 250:1684 (1990)). Several additional members of this family have been discovered but the ligand for erbB2/HER2/neu remains unknown. It is anticipated that further advances in therapeutics will be achieved by the development of therapies that disrupt aberrant growth signaling pathways or affect the cellular interactions of breast cancer cells with native stroma or metastatic sites.
Although oncogenes are likely to be very important in breast cancer, tumor suppressor genes may also play an important role. Certain of these genes, including p53 and Rb-1, are essential to the normal mechanisms that control cell cycle events, especially those checkpoints at the border of the different stages of the cell cycle (Hollstein et al, Science 253:49 (1991); Srivastava et al, Nature 348:747 (1990)).
In 1969, Li and Fraumeni documented a familial cancer syndrome that had an autosomal dominant pattern of expression (Li et al, Ann. Intern. Med. 71:747 (1969)). Members of these families had sarcomas, breast cancers, brain tumors, leukemias, adrenocortical carcinomas, and other malignancies. Family studies demonstrated that the gene responsible for the syndrome was located on chromosome 17, and examination of the p53 gene as a candidate gene revealed that this gene was mutated in five families (Malsin et al, Science 250:1233 (1990)). In the last two years, two genes linked to familial breast cancer, designated BRCA1 and BRCA2, have been isolated and characterized. BRCA1 is at 17q21 (Claus et al, Am. J. Epidemiology 131:961 (1990); Hall et al, Science 250:1684 (1990); Easton et al, Am. J. of Human Genetics 52(4):678 (1993); Black et al, Am. J. of Human Genetics 52(4):702 (1993); Bowcock et al, Am. J. of Human Genetics 52(4):718 (1993); Miki et al, Science 266:66 (1995)). The demonstration of loss of heterozygosity (LOH) at 17q25 has defined another potential tumor suppressor gene (Lindblom et al, Human Genetics 91:6 (1993); Cornelis et al, Oncogene 8:781 (1993); Theile et al, Oncogene 10:439 (1995)).
The present invention relates at least in part, to a novel gene at 17q25 designated K12. K12 encodes a product that is secreted by cancer cells, for example, breast and ovarian cancer cells. The gene is expressed in these tissues at a level that is at least 100 times higher than that of any other malignant or normal tissue examined. The discovery of the K12 gene makes possible novel tumor detection and treatment methodologies.
It is a general object of the invention to provide a novel cancer-related protein and nucleic acid sequence encoding same.
It is another object of the invention to provide a method of detecting the presence of neoplastic cells, including breast and ovarian cancer cells, in a biological sample.
It is a further object of the invention to provide a method of treating a neoplastic condition in a mammal in need of such treatment.
It is yet another object of the invention to provide a method of screening compounds for their ability to bind to, or alter the activity of, the K12 gene product.
In one embodiment, the present invention relates to an isolated nucleic acid encoding the K12 protein, or portion thereof of at least 15 consecutive bases, or complement thereof. The invention also relates to a recombinant molecule comprising such a nucleic acid and a vector, and to a host cell comprising same. In addition, the present invention relates to a method of producing the K12 protein, or portion thereof of at least 5 amino acids, comprising culturing the above-described host cells under conditions such that the nucleic acid is expressed and the K12 protein, or portion thereof, is thereby produced.
In a further embodiment, the present invention relates to an isolated nucleic acid consisting essentially of a double-stranded DNA molecule, one strand of which encodes the K12 protein, or portion thereof of at least 15 consecutive base pairs. The invention also relates to a recombinant molecule comprising such a nucleic acid operably linked to and in inverse orientation with respect to a promoter. The invention further relates to a method of inhibiting K12 protein production in a host cell comprising introducing into the cell such a recombinant molecule under conditions such that the nucleic acid is transcribed and production of the K12 protein is thereby inhibited.
In yet another embodiment, the present invention relates to a method of screening a test compound for its ability to bind to or otherwise alter the growth stimulatory activity of the K12 protein. The method comprises comparing the K12 protein activity using a culture of cells susceptible to the growth stimulatory effects of the K12 protein, in the presence and absence of the test compound. A reduction in the growth of the cells in the presence of the test compound is indicative of a K12 protein inhibitory activity of the test compound, an increase in the growth of the cells being indicative of a K12 protein activating activity of the test compound.
Further objects and advantages of the present invention will be clear from the description that follows.